1. Field of the Invention
The present invention relates to a method for ionizing a substance on a sample, performing mass spectrometry of the substance, and imaging and outputting in-plane distribution of the substance, and an apparatus used therefor.
2. Description of the Related Art
As an analyzing method for comprehensively visualizing distribution information of many substances that constitute body tissue, an imaging mass spectrometry method has been developed, for which a mass spectrometry method is applied. In a mass spectrometry method, a sample is ionized by irradiating a laser light or primary ions and then separated according to a mass-to-charge ratio to obtain a spectrum including the mass-to-charge ratio and detection strength therefor. A sample surface can be subjected to mass spectrometry two-dimensionally so as to obtain two-dimensional distribution of detection strength of a substance corresponding to each mass-to-charge ratio, and obtain distribution information on each substance (mass imaging).
As a mass spectrometry method, a time-of-flight type ion analyzing unit is mainly used that separates and detects ionized target substances depending on differences in time of flight from a sample to a detector. As methods for ionizing the sample, Matrix Assisted Laser Desorption/Ionization (MALDI) of irradiating a pulsed and focused laser light to the sample mixed in a matrix and crystallized, and Secondary Ion Mass Spectrometry (SIMS) of irradiating a primary ion beam to ionize a sample, are known. Among them, the imaging mass spectrometry using MALDI has been widely used for analyzing a biological sample including protein, lipid or the like. However, the MALDI using a matrix crystal limits spatial resolution to several ten μm in principle. Thus, in recent years, Time Of Flight-Secondary Ion Mass Spectrometry (TOF-SIMS), which have high spatial resolution of submicron, has been receiving attention.
In the conventional imaging mass spectrometry method using such methods, a beam for ionization is scanned, and mass spectrometry is successively performed in many minute measurement regions to obtain two-dimensional distribution information. Thus, a considerable time is required to obtain a mass image of a wide region.
To solve this problem, a projection type mass spectrometer has been proposed. In this apparatus, components in a wide region can be collectively ionized, the ions are projected on a detection unit, and thus mass information and two-dimensional distribution of the components can be acquired at one time, thereby measurement time can be significantly reduced. For example, Japanese Patent Application Laid-Open No. 2007-157353 discloses an imaging mass spectrometer that simultaneously records a detection time and a detection position of ions to simultaneously perform mass spectrometry and two-dimensional distribution.
In the time-of-flight mass spectrometer, an axis of an ion optical system that forms a mass spectrometry section is placed perpendicularly to a substrate surface, while generally, a beam for ionization is obliquely incident on a substrate.
When a beam to be a probe is obliquely incident on the substrate, if a substrate or a sample has an irregularity shape (hereinafter referred to as an irregularity on the substrate, or also simply as an irregularity), there appears, around the irregularity, a region to be a shadow to which no beam is irradiated. In this region, a sample is not ionized, and mass spectrometry cannot be performed. Facing this problem, for example, Japanese Patent Application Laid-Open No. 2007-086610 discloses a differential interference microscope including a unit that synthetizes differential interference images obtained from two orthogonal directions, and images a defect with an irregular shape.